Worm Breeder's Gazette 10(3): 7

These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.

Thirty Something

William McCoubrey, Kim Nordstrom and Philip Meneely

In previous Gazettes (WBG 10(1):62-63; 10(2):25-26) we have 
described a series of experiments in which we have used microinjection 
to assay cloned regions of the X-chromosome for feminizing elements.  
Injection of three different regions of the X into tetraploid 
hermaphrodites led to feminization of their chromosomal male progeny 
when those progeny also carried the X-chromosome duplication mnDp8.  
Such animals frequently developed as mosaic intersexes, possessing 
both male and hermaphrodite characteristics.  We are further defining 
the feminizing element(s) using two approaches: reducing the size of 
the injected region and comparing feminizing clones.
A 130 bp sequence consisting of the 3' end of the first intron and 
the first nucleotide of the second exon of act-4 had been shown to 
containing a feminizing element.  Exonuclease III deletions indicated 
that the 5' half of this sequence was not required for feminization.  
An oligonucleotide was synthesized which matched 36 nucleotides at the 
3' end of the intron but did not contain the nucleotide from the exon 
and therefore lacked the splice acceptor site.  When this oligo was 
cloned and tested in our assay, 5 of 7 injected hermaphrodites 
produced a total of 15 intersexual progeny among a total of 82 3A;
2X+Dp animals.  Thus, for this region, the effect has been confined to 
a region of less than 40 bp.
Pairwise comparisons of the three X-chromosomal regions which 
produce intersexes in our assay, act-4, 
a single 8 
nucleotide sequence common to all positive clones.  This sequence, 
TATTGAAA, is present within noncoding regions in all cases.  We are 
testing an oligonucleotide containing only this octamer.  
Interestingly, the identical 8 base sequence is found in the 
regulatory region of X-linked genes in Drosophila melanogaster (E.  
Jaffe, personal communication).  A clone containing this element from 
the white locus of Drosophila (kindly provided by R.  Levis) has been 
tested by the microinjection procedure.  Although only 20 3A;2X+Dp 
progeny arose from nine injected animals, three hermaphrodites 
produced a total of five intersexual progeny.  A clone of the related 
Drosophila autosomal gene, brown (kindly provided by S.  Henikoff), 
did not produce any intersexual animals among 106 duplication-bearing 
males from 18 injected hermaphrodites.  The apparent relationship 
between X-chromosomal sequences from C.  elegans and D.  melanogaster 
is particularly striking because of their great evolutionary 
separation.  Whether there is any functional significance to this 
sequence similarity remains to be explored.